动物模型,慢性心肌缺血 z-bo 2020-08-12 545

　　1. Experimental animals Select specially bred small pigs for experimental use. For chronic experiments, it is best to choose piglets weighing 25-30 kg and about 6 months old to ensure that the operation is convenient and the body weight will not increase too fast at the end of the study.

　　2. Model preparation method: This article mainly introduces the most commonly used Ameroid narrowing ring model preparation method.

　　(1) The preparation of the model should be carried out in an animal operating room with sterile conditions. The operating room should be equipped with anesthesia machine, ECG monitor, defibrillator and other equipment.

　　(2) After the animal is induced to anesthetize, enter the animal operating room, supine position, intubate the trachea and connect the anesthesia machine, and give intravenous anesthesia or inhalation anesthesia during the operation. Continuous ECG monitoring during the operation. If possible, intra-arterial pressure measurement and blood oxygen saturation monitoring can be performed simultaneously.

　　(3) According to the position of the apex beat, a transverse incision is made in the fourth or fifth intercostal space on the left side, and the incision is made layer by layer, and the chest opener assists in exposing the surgical field. Cut the open bag and hang it, you can see the left anterior descending branch and left circumflex branch are located in the anterior interventricular groove and atrioventricular groove respectively. The Ameroid constriction ring is usually placed at the beginning of the circumflex branch, but it can also be placed in the anterior descending branch, but the anterior descending branch should be placed as far as possible after the first larger diagonal branch is issued, about the length of the anterior descending branch. The middle and upper 1/3 junction. Carefully separate the coronary blood vessels at the target site by about 1 cm, and pass the separated blood vessels through the gap of the constriction ring. The constriction ring is fixed around the blood vessel. Suture the pericardium and suture the incision layer by layer. The operation of coronary artery should be minimized during the operation.

　　(4) After the animal is awake, the tracheal intubation is removed. The ECG should be recorded as baseline data after surgery.

　　3. Matters needing attention According to the diameter of the anterior descending vessel, a narrowing ring with an appropriate inner diameter is selected to minimize the possibility of premature occlusion or incomplete occlusion.

　　4. Evaluation of research results

　　(1) Coronary angiography: Coronary angiography is performed about 4 weeks after the operation to determine the degree of target vessel occlusion to confirm whether it meets the requirements of the study.

　　(2) Measurement of myocardial perfusion: The measurement of myocardial perfusion by injection of radioactive microspheres or fluorescent microspheres is a classic evaluation method, which has been widely used in animal experiments related to myocardial ischemia and has a history of many years. The radioactive microspheres are products of Dupont-NEN. There are a variety of radionuclide-labeled microspheres available; the fluorescent microspheres are a set of multiple colors, which can be selected according to research needs. They are produced by Triton (Triton Technology Inc., San Diego CA), can be ordered directly. The injection time of the microspheres can be selected in different ways to achieve different purposes, such as marking the basic perfusion level before the ischemia model is established, or injecting into a specific blood vessel to mark the vascular distribution area, and evaluating the actual situation after modeling. For myocardial perfusion, the same animal can be injected multiple times with different isotope-labeled or different colored microspheres. After the animals were sacrificed, the myocardial tissue was divided according to the research requirements, radioactive tracing or fluorescent microspheres were recovered, the pigments were extracted and analyzed by a fluorescence spectrophotometer to calculate the perfusion of different myocardial tissues. A large number of animal experiments and clinical studies have confirmed that myocardial blood flow is different in different areas of the left ventricle, and the absolute value of myocardial blood flow in a specific area of the ventricle at different times is also different. Therefore, in order to evaluate the changes in regional myocardial perfusion, it is recommended to standardize the data and express the perfusion level of the ischemic area as a percentage of the normal non-ischemic area perfusion, so that it is easy to compare in the series of observations and eliminate the absolute value of myocardial blood flow The impact of the difference in space and time on the results.

　　In live animals, radionuclide myocardial imaging and magnetic resonance imaging can be considered for myocardial perfusion measurement. Under normal circumstances, the cost of these two methods is higher, and they are only considered for application in large animals such as pigs.

　　When using the Ameroid constriction ring model, it must be noted that myocardial ischemia can only occur under load. Therefore, the method of stress test is required. Alternative methods include rapid ventricular pacing (either directly during the operation or implanted in the heart). Outer membrane electrodes for later use), drug load (dopbutamine or dipyridamole, etc.), among which pacing methods are used more and the effect is relatively stable.

　　(3) Evaluation of myocardial function: In miniature pigs, the application of conventional echocardiography can be a good evaluation of the systolic function of the heart. Commonly used indicators include left ventricular ejection fraction (LVEF) to reflect the overall systolic function of the left ventricle, and left ventricular systolic wall thickening rate (WT%) to reflect the systolic function of a certain part of the left ventricle. If radionuclide myocardial imaging is performed for animals, the function of the myocardium can be evaluated at the same time.

　　(4) Histology: Microangiogenesis and arteriogenesis (angiogenesis and arteriogenesis) should be evaluated by histological methods, and immunohistochemical staining with specific markers of vascular endothelial cells and vascular smooth muscle cells should be used to confirm the presence of vascular structures. Under a microscope count. Commonly used vascular endothelial cell markers include CD31, vWF, and vascular smooth muscle cells are usually labeled with α-actin.